HIV-1 DNA vaccines induce both humoral and cellular immune responses in experimental animals, suggesting their potential usefulness for immunization. However, further optimization of DNA vaccines is needed to induce protection against virulent HIV-1 isolates. We propose to utilize CD40 activation and targeting in an attempt to improve the ability of HIV-1 env DNA vaccines to generate strong cellular immunity and high titers of neutralizing antibodies. We will construct DNA vaccines encoding CD40 ligand (CD154) or a single chain Fv (scFv) specific for CD40, fused with DNA encoding portions of the HIV-1 env protein. After DNA vaccination, the expression of this fusion protein in vivo could result in both activation of the CD40 receptor and direction of HIV-1 env antigens into the endocytic pathway of CD40 positive antigen presenting cells (APC). Internalization of env antigens after binding the CD40 positive antigens presenting cells (APC). Internalization of env antigens after binding the CD40 receptor will enhance presentation of peptides to effective antibody production and generation of CD4+ helper T cell and CD8+CTL activity. We hypothesize that the combination of CD40 activation and presentation of viral peptides by MHC class II will lead to improved immune responses to viral antigens. This hypothesis will be tested by construction of anti-human CD40 scFv-env DNA vaccines, followed by testing in primates where the anti-human CD40 scFv and human CD154 retain CD40 binding and functional activity. Antibody responses towards HIV-1 env will be tested by epitope specific ELISA assays and viral neutralization assays.